100 results about "Differential centrifugation" patented technology

The invention discloses an implant-derived exosome which is prepared by the following steps: adding cleaned and crushed colored rice flour into a PBS (Phosphate Buffer Solution) with the pH value of 6-7.4, homogenizing at a high speed, adding alpha-amylase into rice milk, performing enzymolysis for 1-2 hours under the conditions that the pH value is 6-7.4 and the temperature is 90-97 DEG C, filtering, and standing filtrate overnight; carrying out gradient differential centrifugation at 4 DEG C, collecting supernate, filtering, carrying out ultracentrifugation on filtrate at 4 DEG C and 12000-15000g for 1.5-3 hours, and collecting precipitate; and re-suspending the precipitate, enriching by using an MWCO ultrafiltration tube with the molecular weight cut-off of 100kDa, collecting the precipitate at the bottom of the ultrafiltration tube, and re-suspending in a PBS buffer solution, thereby obtaining the product. The exosome can effectively inhibit the generation of inflammatory factors, has a good inhibition effect on various pathogenic bacteria, also has an anti-oxidation effect, and has application prospects in the fields of inflammation resistance, bacteria resistance, oxidation resistance and the like.

The invention belongs to the field of biotechnology and aquaculture, and particularly relates to a method for improving hypoxia resistance of pelteobagrus vachelli based on hypoxia induction of liver extracellular vesicles of the pelteobagrus vachelli. The method comprises the following steps: extracting high-purity extracellular vesicles EVs in liver tissues of pelteobagrus vachelli subjected to low-oxygen treatment by adopting a method of combining differential centrifugation and high-precision iodixanol density gradient centrifugation; diluting the extracted EVs to 2mu g / mu L by using PBS (Phosphate Buffer Solution) to prepare an EVs suspension; an intraperitoneal injection mode is adopted, the EVs suspension is injected into the body of the pelteobagrus vachelli, and after injection is conducted for 24 hours, the hypoxia resistance of the pelteobagrus vachelli is evaluated through a suffocation point detection test. The invention opens up a brand new path for improving the hypoxia resistance of the pelteobagrus vachelli, provides a new idea and a new method for efficiently enhancing the hypoxia resistance of economic fishes in the field of aquaculture, is expected to play an important role in actual aquaculture production, and assists the sustainable development of the aquaculture industry.

The invention relates to an astragaloside-loaded astragalus membranaceus exosome which is prepared by the following steps: crushing astragalus membranaceus, and filtering to obtain juice; performing multi-step differential centrifugation to obtain supernate; carrying out ultrafiltration and centrifugation, and collecting the precipitate; adding to the upper layer of a sucrose stepped gradient solution, centrifuging, and collecting part of strips; after dilution, centrifuging, discarding the supernatant, precipitating, and removing cane sugar to obtain the astragalus membranaceus exosome; the radix astragali exosome loaded with the astragaloside is obtained by mixing the radix astragali exosome with the astragaloside and carrying out ultrasonic treatment or co-incubation, and can be used for preparing medicines for specifically inhibiting cGAS / STING pathway excessive activation in cerebral ischemia reperfusion injury and downstream NLRP3 inflammasome mediated pyroptosis and preparing medicines for protecting and repairing mitochondria in the cerebral ischemia reperfusion injury. The homologous astragalus membranaceus exosome is used as a natural carrier, the blood-brain barrier is effectively overcome, and the enrichment concentration and bioavailability of astragaloside in a cerebral ischemia area are remarkably improved.

The invention discloses a preparation method of vascular tissue-derived extracellular vesicles, and belongs to the field of cytobiology. According to the preparation method of the extracellular vesicles, the artery blood vessel tissue is directly subjected to enzymolysis through collagenase, the extracellular vesicles are obtained by combining low-speed centrifugation and ultra-speed centrifugation methods and utilizing a differential centrifugation mode, and the technical blank of obtaining the extracellular vesicles from the blood vessel tissue is filled. Experimental results show that the extracellular vesicles can be successfully prepared from collagenase I, collagenase II, collagenase IV, collagenase V or collagenase IV combined with elastin, the prepared extracellular vesicles are different in particle size, form, yield, protein quantity, purity and the like, and the quality of the extracellular vesicles obtained by enzymolysis of collagenase IV combined with elastin is optimal. The extracellular vesicles are extracted from vascular tissues with normal physiological characteristics for the first time, a new source is provided for preparation of the extracellular vesicles, and the extracellular vesicles have great significance in expanding application of the extracellular vesicles.

The invention discloses an immune agonist and a preparation method thereof, and relates to the technical field of immune agonist preparation, and the preparation method of the immune agonist comprises the following specific steps: S1, preparing a double-target immune agonist monomer; s2, preparing a cationic liposome inner core; s3, biomimetic cell membrane coating; and S4, finished product detection. According to the immune agonist and the preparation method thereof, in the preparation process, a continuous flow microreactor is used for two-step synthesis of a double-target monomer, a segmented precise temperature control mode is matched with microwave strengthening, the reaction period can be shortened, efficient entrapment of the agonist is achieved through a micro-reaction technology, and the problems that a traditional batch method is low in entrapment efficiency and uneven in particle size are solved; in addition, MDC cell membrane extraction is conducted through differential centrifugation, the integrity of the membrane is guaranteed, active targeting of a CD11c marker on the surface of the MDC cell membrane is achieved through a cell biomimetic membrane, and after lipidosome and membrane vesicles with the volume ratio being 3: 1 are fused, the in-vitro BMDC binding rate is increased compared with that of naked lipidosome.

The invention discloses a composition containing ginseng exosome and super-elastic protein and application of the composition, and belongs to the technical field of biology. The composition containing the ginseng exosome and the super-elastic protein is prepared by compounding the ginseng exosome and the super-elastic protein according to the weight ratio of 10: (100-150), the ginseng exosome is prepared by the following method: extracting the ginseng exosome from ginseng root tissues by using a differential centrifugation method to obtain a ginseng exosome crude extract; adding sucrose solutions with different concentrations into the ginseng exosome crude extract, and performing ultra-high-speed centrifugal purification to obtain purified ginseng exosome; the amino acid sequence of the super-elastic protein is as shown in SEQ ID NO. 3. According to the invention, the ginseng exosome and the super-elastic protein are combined for use for the first time, and both the ginseng exosome and the super-elastic protein have the effect of promoting cell proliferation, so that the use safety is high; the ginseng exosome and the super-elastic protein can synergistically improve the skin tightening and anti-wrinkle effects and the skin injury repairing effect.

The invention belongs to the technical field of thermal interface materials, and particularly relates to a boron nitride-based flexible heat-conducting composite film and a preparation method and application thereof.The preparation method comprises the following steps that h-BN powder and water are mixed and subjected to ball milling, centrifugation is conducted after ball milling, supernate is taken, and BNNS dispersion liquid is obtained; carrying out differential centrifugation on the obtained BNNS dispersion liquid to obtain large-sheet-diameter BNNS, and carrying out uniform ultrasonic dispersion on the large-sheet-diameter BNNS in a differential alcohol organic solvent to obtain large-sheet-layer BNNS organic dispersion liquid; the preparation method comprises the following steps: performing oxidation modification on cellulose nanofibers by adopting a TEMPO oxidation method, and uniformly dispersing the oxidized and modified cellulose nanofibers in water to obtain an aqueous dispersion of the oxidized and modified CNF; the preparation method comprises the following steps: mixing an organic dispersion liquid of large-lamellar BNNS with an aqueous dispersion liquid of oxidation modified CNF, firstly stirring, and then carrying out ultrasonic mixing to obtain a BNNS / CNF composite dispersion liquid; and carrying out suction filtration on the BNNS / CNF composite dispersion liquid to obtain a wet film, and drying and tabletting the wet film to obtain the boron nitride-based flexible heat-conducting composite film. The preparation method is suitable for continuous and large-scale preparation of the high-performance boron nitride-based composite heat-conducting film.

The utility model relates to the technical field of water quality monitoring, and discloses a differential centrifugal device for water quality monitoring, which comprises a bottom plate and a mounting shell, and further comprises a clamping assembly arranged in a containing cavity and used for increasing the loading and unloading speed of a centrifugal tube, the driving assembly is arranged on one side of the mounting shell and can provide rotating force for the transmission shaft. According to the utility model, the synchronous positioning of the centrifugal tubes in the plurality of placing grooves can be realized through the clamping assembly, so that the loading and unloading process of the centrifugal tubes is simplified, the loading and unloading difficulty of the centrifugal tubes is reduced, and the labor burden of detection personnel is relieved; the rotating drum is driven by the driving assembly to rotate to separate and precipitate different substance particles in a water sample, the turbidity test of a water body is completed, the conditions of distortion, breakage and the like of the transmission shaft due to overlarge centrifugal force can be avoided through the limiting assembly, and the stability of the structure is improved.

This invention belongs to the field of biomedical technology, specifically relating to a madder-derived exosome-like nanovesicle, an artificial blood vessel, and their preparation and application. This invention utilizes differential centrifugation and sucrose density gradient centrifugation techniques to extract high-purity, highly active exosome-like nanovesicles from madder. Then, polydopamine is loaded onto the surface of a small-diameter polyurethane artificial blood vessel, which is then immersed in a suspension of madder-derived exosome-like nanovesicles to obtain a small-diameter artificial blood vessel loaded with madder-derived exosome-like nanovesicles. These madder-derived exosome-like nanovesicles can effectively promote endothelial cell proliferation and inhibit reactive oxygen species generation; animal experiments show unobstructed blood flow under ultrasound. The small-diameter artificial blood vessel loaded with madder-derived exosome-like nanovesicles can simultaneously achieve in vivo endothelialization and vascular patency, possessing significant clinical application value.

The invention relates to the technical field of biological medicine, and provides application of a dental pulp stem cell exosome in preparation of a medicine for treating and preventing osteoarthritis, a preparation method of the dental pulp stem cell exosome comprises the following steps: S1, obtaining dental pulp tissue from a dental pulp cavity of an in-vitro tooth; s2, pretreating dental pulp tissues, adding digestive juice, digesting, centrifuging, discarding supernatant, adding a complete culture medium containing antibiotics into precipitates, mixing, filtering and centrifuging to obtain dental pulp stem cells; and S3, culturing the dental pulp stem cells by using a complete culture medium, when the confluence degree of the dental pulp stem cells reaches 60-80%, replacing the serum-free culture medium for continuous culture, collecting the dental pulp stem cell conditioned culture medium, and performing differential centrifugation to obtain the dental pulp stem cell exosome. According to the technical scheme, the regulating effect of the dental pulp stem cell exosome on endoplasmic reticulum stress and the mechanism of the dental pulp stem cell exosome for improving cartilage cell degeneration are explored.

The application belongs to the technical field of cell biology, and particularly relates to a solution rich in Sargassum fusiforme nano-vesicles and a preparation method and application thereof. The solution rich in Sargassum fusiforme nano-vesicles is prepared by a differential centrifugation combined with hollow fiber membrane concentration, and the obtained solution rich in Sargassum fusiforme nano-vesicles can play an anti-inflammatory role, inhibit high-fat diet induced fatty liver and osteoporosis, and can also be used for preparing a mask with soothing, moisturizing and anti-wrinkle effects.

The invention discloses application of exosome-like nanoparticles derived from fructus lycii to preparation of a medicine for preventing or treating oligospermia and / or asthenospermia. Sterile, stable and high-purity fructus lycii exosome-like nanoparticles (LBELNs) are obtained by adopting a differential centrifugation-suction filtration-ultra-high-speed centrifugation method, the physiological efficacy of the LBELNs is researched, and it is found that the LBELNs can significantly improve the oligoasthenospermia disease activity index of male mice induced by cyclophosphamide (CTX); the sperm quantity is obviously increased, the sperm malformation is improved, the sperm motility is improved, and the sex hormone level is recovered. The LBELNs can effectively relieve spermatogenic dysfunction of testicular tissues, the sperm differentiation and formation process is obviously improved, the total number of sperms is increased, and the treatment effect is more obvious along with increase of the dosage of the LBELNs. The invention has potential in preparation of drugs for preventing or treating oligoasthenospermia, and provides a new strategy and treatment method for treating oligoasthenospermia.

The invention relates to the technical field of medicines, in particular to an extraction method and application of cordate houttuynia exosome-like nanoparticles. The method comprises the steps of pretreatment, impurity removal through differential centrifugation, fine filtration, ultra-speed centrifugation crude extraction, sucrose density gradient centrifugation purification, washing and the like. By optimizing key process parameters, the HELNs with the average particle size of 50-150nm and the typical double-layer membrane cup-shaped vesicle form can be stably obtained. Experiments prove that the HELNs prepared by the method have independent and direct anti-staphylococcus aureus activity. Based on the newly discovered biological activity, the invention provides new application of the HELNs in preparation of products (such as antibacterial drugs, dressings, coatings, lotions and the like) for inhibiting staphylococcus aureus and / or treating infection of staphylococcus aureus. The method is standard in operation and good in repeatability, and widens the application field of active ingredients of houttuynia cordata.

The invention discloses a stem cell-based microvesicle as well as a preparation method and application thereof, belongs to the technical field of biological tissue engineering, and particularly relates to a preparation method of the stem cell-based microvesicle, which comprises the following steps: culturing stem cells until the cell fusion degree is 80-90%, performing hypoxia treatment, and cleaning for 2-3 times; standing and culturing the cleaned stem cells, then adding a calcium ion carrier, and carrying out ultrasonic treatment; the serum-free basic culture medium containing the calcium ion carrier is immediately absorbed and removed, the cells are cleaned through PBS, culture is conducted for 24 h, and cell supernatant is collected and merged respectively; performing first centrifugation to remove cell debris; performing second centrifugation to remove dead cells and large cell debris; carrying out third centrifugation to remove organelles; fourth centrifugation is carried out, large vesicles are removed, and a final supernatant is obtained; and filtering and centrifuging to obtain a microvesicle precipitate. The preparation method of the microvesicles aims at systematically improving the yield and biological activity of the stem cell microvesicles through low-oxygen pre-stimulation, ultrasonic-calcium ion carrier synergistic induction and differential centrifugation.

The invention discloses a preparation method for industrial production of plant-derived exosomes, and relates to the technical field of biological product preparation. Comprising the following steps: S1, pretreatment of raw materials: cleaning and disinfecting plant tissues, mixing the plant tissues with a buffer solution, and homogenizing to obtain a plant crude extract; s2, pre-clarification: sequentially carrying out differential centrifugation and step-by-step filtration on the crude extract obtained in the step S1 to remove large-particle impurities so as to obtain pre-clarified liquid; s3, tangential flow filtration: performing concentration and buffer solution replacement on the pre-clarified solution through a tangential flow filtration system to obtain a primarily purified exosome concentrated solution. According to the method disclosed by the invention, industrial, continuous and scalable production of the plant exosome is realized, the product purity is high, and the batch difference is small; impurities influencing gelation are removed through an optimized purification process, a method for compounding the exosome with polysaccharide hydrogel under a low shear condition is defined, the formed compound preparation improves the structural stability of the exosome by 2-5 times, and local sustained-release administration is realized.

This invention belongs to the field of ultrasound contrast microbubble synthesis technology, and relates to a method for preparing and applying ultrasound contrast microbubbles targeting CD93 molecules in tumor vascular endothelium. The method includes the following steps: constructing a recombinant expression vector capable of expressing the CD93 ligand MMRN2 protein, transfecting cells, and extracting the cell membrane using differential centrifugation; mixing dipalmitoylphosphatidylcholine, distearate phosphatidylethanolamine-polyethylene glycol 2000, and chloroform, adding DiI, and preparing a liposome membrane using a thin-film hydration method; mixing the cell membrane and the liposome membrane, performing ice-bath sonication, and mixing and shaking with perfluoropropane to obtain an ultrasound contrast agent targeting CD93 molecules in tumor vascular endothelium. The ultrasound contrast microbubbles prepared by the method of this invention have good biocompatibility compared to conventional antibody-targeting strategies, and can also improve stability and targeting, thereby enabling the assessment of CD93 molecule expression in tumors.

The invention discloses an exosome composition containing resveratrol, astragalus membranaceus and ginseng as well as a preparation method and application of the exosome composition, and belongs to the technical field of biological medicines. The exosome composition disclosed by the invention is obtained by inducing alveolar epithelial cells by active ingredients containing resveratrol, an astragalus extract and a ginseng extract and then secreting the alveolar epithelial cells. The preparation method of the exosome composition mainly comprises the following steps: co-culturing the active component and alveolar epithelial cells, and then sequentially carrying out differential centrifugation and ultracentrifugation on the collected cell supernatant to purify the exosome. The invention also provides a pharmaceutical preparation containing the exosome composition. According to the composition, traditional Chinese medicine active ingredients are combined with a modern cell exosome technology, a potential strategy is provided for prevention and treatment of pulmonary fibrosis, and the composition has a good application prospect.

The invention relates to a method for improving secretion and quality of stem cell exosomes, and belongs to the technical field of cell culture.The method specifically comprises the following steps that S1, stem cells are resuspended in a basic culture medium and subjected to resuscitation treatment, and pretreated stem cells are obtained; s2, taking a basic culture medium, removing FBS of exosomes, and then adding ergothioneine and beta-nicotinamide mononucleotide according to the final concentration to obtain an improved culture medium; and S3, washing the pretreated stem cells, adding an improved culture medium, culturing, carrying out differential centrifugal precipitation on the exosome, and filtering and purifying through a filter membrane. According to the technical scheme, the ergothioneine and the NMN serve as components of an improved culture medium at the same time, ROS is removed by the ergothioneine, degradation of NAD < + > by the ergothioneine can be reduced, the NMN provides ATP support for the antioxidant process of the ergothioneine, and the ergothioneine and the NMN cooperate to improve the exosome secretion quality.

The application relates to a method for purifying umbilical cord blood mesenchymal stem cell-derived exosomes, and relates to the technical field of bioengineering. The method combines gradient centrifugation and membrane filtration. First, the cell culture supernatant is pretreated by adjusting the pH, and cell debris and large-particle impurities are removed by multi-stage differential centrifugation. Then, selective protein precipitation is carried out by using polyethylene glycol with a specific molecular weight, and the exosomes are finely separated by sucrose-meglumine diatrizoate density gradient centrifugation. Finally, terminal purification is completed under low-temperature and low-pressure conditions by using a polyether sulfone membrane filtration system with optimized pore size. By precisely controlling the centrifugation parameters, reagent concentration and filtration conditions, the exosomes obtained by the method have a particle size concentrated in the range of 30-150 nm, a protein purity of more than 95%, and a recovery rate of more than 85%, and the separation efficiency and quality of the exosomes are significantly improved.

The invention discloses a bifidobacterium breve extracellular vesicle as well as a preparation method and application thereof, and belongs to the field of biological medicines. The extracellular vesicles are obtained by the following steps: carrying out anaerobic culture on Bifidobacterium breve FB-342 in a culture medium containing L-cysteine, and then carrying out differential centrifugation, filtration and ultracentrifugation on the cultured Bifidobacterium breve FB-342. According to the invention, the extracellular vesicles with definite physicochemical characteristics (the average particle size is about 143.1 nm and the purity is about 1.76 * 10 < 11 > parts / g protein) are successfully prepared from the bifidobacterium breve FB-342 for the first time, and experimental results show that the prepared extracellular vesicles of the bifidobacterium breve FB-342 have obvious potential for treating ulcerative colitis in cell and animal models, and can be used for preparing the extracellular vesicles of the bifidobacterium breve FB-342 for treating ulcerative colitis. And a new strategy based on the probiotic vesicles is provided for treatment of ulcerative colitis.

The invention discloses an extraction method of hawthorn exosomes and application of the hawthorn exosomes in treatment of DSS-induced colitis, and belongs to the technical field of plant exosomes. The extraction method comprises the steps of raw material treatment, differential centrifugation, membrane filtration, ultracentrifugation enrichment, density gradient purification and desugaring refining, and the high-purity crataegus pinnatifida exosome with the capsule-shaped structure can be obtained. The hawthorn exosome intervenes DSS-induced colitis through preventive intragastric administration (25-50 mg / kg / d), the disease activity index (DAI) can be reduced to 2 or below, the expression quantity of colon tissue TNF-alpha is recovered to the normal level, the IL-1beta inhibition effect is superior to that of a hawthorn extract, and the curative effect is remarkably superior to that of plant exosomes such as rhizoma curcumae longae, tea leaves and black tea. The process is standardized, the curative effect is definite, and a new scheme is provided for colitis treatment.

This invention discloses an umbilical cord mesenchymal stem cell membrane for treating hair loss, its preparation method, and its application, belonging to the field of biomedical technology. The preparation method of the umbilical cord mesenchymal stem cell membrane includes the following steps: obtaining human umbilical cord mesenchymal stem cells, digesting, centrifuging, washing, and resuspending them to obtain a resuspended cell solution; subjecting the resuspended cell solution to repeated freeze-thaw cycles to break the cells, obtaining a cell lysate; and subjecting the cell lysate to differential centrifugation to obtain the umbilical cord mesenchymal stem cell membrane. This umbilical cord mesenchymal stem cell membrane retains the natural cell membrane structure and functional proteins, has a negatively charged surface, and exhibits good dispersibility and stability. It can improve the survival rate of dermal papilla cells without introducing live cells and effectively regulate the skin microecology and metabolite balance, demonstrating high safety, good biocompatibility, and excellent hair growth promotion effects.

The invention discloses waste cigarette cut tobacco recovery equipment, belongs to the field of auxiliary equipment for tobacco processing, and aims to solve the problems of disordered cigarette conveying, low cut tobacco removal accuracy, insufficient resource recovery and the like of existing equipment. The equipment comprises a centrifugal disc device and a cigarette tobacco shred recovery device, the centrifugal disc arranges scattered cigarettes through differential centrifugation, the recovery device comprises a rack, a longitudinal / transverse conveying assembly, a tobacco shred removing assembly, a staggered conveying assembly, a spray gun, a tobacco shred conveyor and other assemblies, and ordered cigarette conveying, directional tobacco shred removing and classified recovery can be achieved. Stable conveying is guaranteed through a guide limiting structure, tobacco shreds, filter tips and cigarette paper are accurately separated through a spray gun assembly, qualified cigarettes and unqualified cigarettes are separated through the double-conveying-belt design, and classified recycling is achieved through multiple collecting boxes. The equipment adapts to cigarettes of different specifications, runs stably, is convenient to operate, can improve the tobacco shred recovery efficiency and purity, and reduces resource waste and recovery cost.

The invention relates to an exosome composition for treating insomnia and a preparation method thereof. In one aspect, the present invention relates to the use of a combination of a ginseng exosome and a stem cell exosome in the preparation of a medicament for the treatment or prevention of insomnia. The ginseng exosome is prepared by a method comprising the following steps: taking fresh ginseng roots, adding a PBS (Phosphate Buffer Solution) for homogenizing, and filtering to obtain a ginseng solution; extracting the ginseng solution by using a differential centrifugation method to prepare an exosome suspension; the invention also relates to an exosome composition prepared by mixing the ginseng exosome and the stem cell exosome. The invention further relates to a preparation method of the exosome composition. The exosome can be used for treating or preventing insomnia.

The invention provides a preparation method of extracellular vesicles carrying mycobacterium tuberculosis specific antigens, and belongs to the technical field of extracellular vesicles. THP-1 mononuclear cells are induced to be differentiated into macrophages through phorbol ester, a mycobacterium tuberculosis H37Rv standard strain is treated through ultrasonication and needle blowing and beating to prepare a monodisperse bacterium solution, and the monodisperse bacterium solution is added into the mycobacterium tuberculosis specific antigens to prepare the extracellular vesicles carrying the mycobacterium tuberculosis specific antigens. Infecting according to the infection complex value of 5, monitoring the real-time infection rate every 12 hours in the culture process, dynamically adjusting the initial bacterial liquid concentration according to the value to maintain the infection rate in the range of 45-55%, purifying the extracellular vesicles through a differential centrifugation method, detecting the particle size and concentration of the vesicles and the content of lipoarabinomannan, and determining the content of lipoarabinomannan in the extracellular vesicles. Qualified vesicle products are divided into three concentration grades according to the antigen content, and dimethyl sulfoxide is added for cryopreservation, so that the technical problem of poor antigen loading stability among batches of extracellular vesicle products carrying mycobacterium tuberculosis specific antigens is solved.

The invention provides pogostemon cablin extracellular vesicles and an extraction and separation method and application thereof. Pogostemon cablin tissue juice is subjected to differential centrifugation, PEG precipitation and ultrafiltration, and the pogostemon cablin extracellular vesicles are extracted. The method is simple and convenient, equipment and materials are easy to obtain, the obtained extracellular vesicle crude extract is purified by combining an ultrafiltration method, and the obtained purified solution is clear and transparent and high in concentration and purity. The compound has no toxicity to HaCaT human immortalized epidermal cells and zebrafish, has good biocompatibility, shows good anti-photoaging, anti-oxidation and anti-inflammatory effects in in-vitro and in-vivo experiments, and is expected to be prepared into an anti-photoaging natural plant drug.

This invention relates to the field of biotechnology, specifically to nanoparticles prepared from *Hedyotis diffusa* and their applications. The invention uses *Hedyotis diffusa* as raw material and prepares nanoparticles by differential centrifugation at low, medium, and ultra-high speeds. The nanoparticles have a particle size of 100-200 nm, exhibit a spherical membrane structure, and are uniformly distributed. These nanoparticles contain various proteins, lipids, and RNA. Oral delivery of *Hedyotis diffusa*-derived nanoparticles effectively activates immune cells in the intestine, including T cells, especially γδT cells. Experiments have shown that these *Hedyotis diffusa*-derived nanoparticles can significantly reverse the immunosuppressive tumor microenvironment of colon cancer in the intestine and delay tumor growth.

The invention discloses a preparation method and application of ganoderma lucidum outer vesicles without sporocarp waste sources. The ganoderma lucidum external vesicles are used for preparing medicines for treating ischemic cerebral apoplexy sequelae, and can improve core symptoms such as dyskinesia (including limb paralysis, imbalance and gait abnormity), cognitive decline and the like after apoplexy. The spore-free sporocarp waste sourced ganoderma lucidum outer vesicles are prepared from spore-free sporocarp waste which is completely released by spores after a ganoderma lucidum powder spraying period through raw material pretreatment, differential centrifugation, tangential flow concentration and filter membrane filtration. According to the preparation method, the waste is used as a raw material to prepare the high-value medical outer vesicle, green circulation of agricultural waste-high-added-value medical raw materials is achieved, the ganoderma lucidum outer vesicle can effectively penetrate through the blood-brain barrier, brain-targeted delivery is achieved, and the problems that traditional ganoderma lucidum active ingredients are low in blood-brain barrier penetration rate and poor in blood-brain barrier penetration rate are solved. The effective treatment concentration in the brain tissue is difficult to achieve, and the traditional drug delivery bottleneck is broken through.

The invention discloses a preparation method and application of a mesenchymal stem cell exosome which is enriched with Foxp1 protein and has immune organ targeting property. The method comprises the following steps: acquiring a single-cell suspension of the mesenchymal stem cells, performing suspension culture on the single-cell suspension of the mesenchymal stem cells in an exosome removal culture medium to induce the mesenchymal stem cells to aggregate to form a compact cell cluster, and then collecting the exosome through differential centrifugation. According to the method disclosed by the invention, the mesenchymal stem cells are induced to aggregate through a suspension culture technology, so that the exosome with high yield and immune targeting is obtained. The Foxp1 protein is specifically enriched in the exosome, the exosome has the characteristics of high yield, immune organ targeting and strong immune regulation capability, Treg cell differentiation is regulated through the Foxp1 / STAT5 / Foxp3 axis, the limitation in the application of the existing exosome is solved, and a new strategy is provided for the treatment of autoimmune diseases.

The application discloses a method for extracting mitochondria-associated membranes (MAM) and other subcellular components from podocytes, which comprises the following steps: pretreating the podocytes, crushing the podocytes by low-power ultrasonic, separating the crude mitochondrial components by differential centrifugation, and then purifying the high-purity MAM and other subcellular components by density gradient centrifugation and ultracentrifugation. The method replaces the traditional homogenizer grinding with low-power ultrasonic crushing, improves the MAM extraction yield by controlling the ultrasonic parameters, appropriately simplifies the differential centrifugation steps, increases the yield while ensuring the purity, sequentially separates the unbroken cells, the crude mitochondria, the cytoplasm and the endoplasmic reticulum components, and finally obtains the high-purity MAM and other subcellular components by density gradient centrifugation and ultracentrifugation purification. The method effectively improves the yield of the podocyte separation and extraction of the MAM and other subcellular components, reduces the cost, and provides technical support for the research on the podocyte MAM proteomics.